PROJECT SUMMARY Brain tumors have become the leading cause of cancer-related death in children. Ependymoma (EPN) accounts for a substantial number of these deaths. Over 70% of children presenting with ependymoma will relapse and almost all children who relapse will eventually die. Unlike medulloblastoma, no effective chemotherapy has been developed in ependymoma to complement the standard, and eventually for most ineffectual, surgery and radiation. In particular, children with the most common type of ependymoma, posterior fossa Group A (PFA1), relapse more frequently and experience more invasive, metastatic disease at relapse. Thus, there is a critical need for more effective therapies to combat high-risk PFA1 tumors. Single-cell RNAsequencing data suggest that the epigenetic silencing of LDOC1 within a specific subpopulation of tumor cells has a profound, direct impact on the tumor biology of PFA1 tumors. The driving hypothesis is that epigenetic silencing of LDOC1 with in the MEC subpopulation, as a result of chromatin remodeling, is the molecular driver in PFA1 EPN, through upregulation of non-canoncial NF-?B activation. To address this hypothesis, the studies in aim one will determine the role of LDOC1 expression in EPN by examining 1) the mechanism of gene silencing, 2) the functional role of loss of LDOC1 in vitro and in vivo, and 3) the genomic transcriptional targets of LDOC1. Aim two is designed to 1) determine how LDOC1 regulates non-canoncial NF-?B signaling and 2) identify the functional consequences of the NF-?B signaling pathway. The collective proposed studies will define the effect of LDOC1 loss, which we hypothesize to be the molecular driver of tumor biology of PFA1 EPN. These studies will significantly add to our understanding of childhood EPN and have the potential to identify rational therapeutic targets for children with this high-risk, poor-outcome pediatric brain tumor.